Synergistic neutrophil elastase-cytokine interaction degrades collagen in three-dimensional culture

Proteolytic degradation of extracellular matrix is thought to play an important role in many lung disorders. In the current study, human lung fibroblasts were cast into type I collagen gels and floated in medium containing elastase, cytomix (combination of tumor necrosis factor-alpha, interleukin-1beta, and interferon-gamma), or both. After 5 days, gel collagen content was determined by measuring hydroxyproline. Elastase alone did not result in collagen degradation, but in the presence of fibroblasts, elastase reduced hydroxyproline content to 75.2% (P < 0.01), whereas cytomix alone resulted in reduction of hydroxyproline content to 93% (P < 0.05). The combination of elastase and cytomix reduced hydroxyproline content to 5.2% (P < 0.01). alpha(1)-Proteinase inhibitor blocked this synergy. Gelatin zymography and Western blot revealed that matrix metalloproteinase (MMP)-1, -3, and -9 were induced by cytomix and activated in the presence of elastase. Tissue inhibitor of metalloproteinase (TIMP)-1 and -2 were also induced by cytomix but were cleaved by elastase. We conclude that a synergistic interaction between cytomix and elastase, mediated through cytokine induction of MMP production and elastase-induced activation of latent MMPs and degradation of TIMPs, can result in a dramatic augmentation of collagen degradation. These findings support the notion that interaction among inflammatory mediators secreted by mononuclear cells and neutrophils can induce tissue cells to degrade extracellular matrix. Such a mechanism may contribute to the protease-anti-protease imbalance in emphysema.     

Diverse repertoire of the MHC class II-peptide complexes is required for presentation of viral superantigens

Among other features, peptides affect MHC class II molecules, causing changes in the binding of bacterial superantigens (b-Sag). Whether peptides can alter binding of viral superantigens (v-Sag) to MHC class II was not known. Here we addressed the question of whether mutations limiting the diversity of peptides bound by the MHC class II molecules influenced the presentation of v-Sag and, subsequently, the life cycle of the mouse mammary tumor virus (MMTV). T cells reactive to v-Sag were found in mice lacking DM molecules as well as in A(b)Ep-transgenic mice in which MHC class II binding grooves were predominantly occupied by an invariant chain fragment or Ealpha(52-68) peptide, respectively. APCs from the mutant mice failed to present v-Sag, as determined by the lack of Sag-specific T cell activation, Sag-induced T cell deletion, and by the aborted MMTV infection. In contrast, mice that express I-A(b) with a variety of bound peptides presented v-Sag and were susceptible to MMTV infection. Comparison of v-Sag and b-Sag presentation by the same mutant cells suggested that presentation of v-Sag had requirements similar to that for presentation of toxic shock syndrome toxin-1. Thus, MHC class II peptide repertoire is critical for recognition of v-Sag by the T cells and affects the outcome of infection with a retrovirus.     

The psychologic consequences of exposure to high density pulsed electromagnetic energy

Five albino female rats which had been trained to run a maze were exposed to 3 nanosecond pulses of electromagnetic energy which had a density of 600,000 v/m². A disturbance of the ability of the animals to perform this recently learned task was observed.The effect was found to be reversible within a period of 30 min. Possible mechanisms by which the decision making strategy of the animals was temporarily disrupted are considered. The parallelism between the conditions of this experiment and those attending the exposure of animals to lightning bolts during electrical storms is pointed out.

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Zusammenfassung Fünf weibliche Albinoratten, die darauf trainiert waren, in einem Labyrinth zu laufen, wurden 3 Nanosekunden-Impulsen elektromagnetischer Energie ausgesetzt, die eine Energie von 600.000 v/m² hatten. Es wurde eine Störung der Fähigkeit der Tiere beobachtet, diese kürzlich gelernte Aufgabe zu bewältigen. Die Wirkung war innerhalb von 30 Min. reversibel. Die verschiedenen Mechanismen, durch die die Fähigkeit der Tiere, ihre Aufgabe zu lösen, zeitweise unterbunden wurde, werden diskutiert. Auf die Parallele zwischen den Bedingungen dieses Experiments und denen bei Entladungen während elektrischer Stürme wird hingewiesen.

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Resume On a exposé cinq femelles de rats albinos à des impulsions électromagnétiques de 3 manosecondes et d'une énergie de 600.000 v/m². Ces bêtes avaient au préalable été entraînées à se mouvoir dans un labyrinthe.On a alors pu constater que les bêtes ainsi exposées perdaient la faculté de se retrouver dans le labyrinthe. L'effet de l'exposition est cependant réversible après 30 minutes.On discute les divers mécanismes par lesquels la faculté qu'ont les bêtes d'effectuer certaines tâches est momentanément supprimée. On mentionne le parallélisme existant entre les conditions de ces expériences et celles que provoquent les décharges qui se produisent pendant les orages.

     

Incidence of phenylketonuria in British Columbia, 1950-1971

The incidence of PKU in British Columbia in the 1950-1971 period is 1/18,750 which corresponds to that found in two other Canadian studies.^{2, 3} Evidence is presented which shows a trend toward a decline in incidence; however, this is not statistically significant. There is a preponderance of male cases in all age groups.     

Endohexosaminidase-catalysed glycosylation with oxazoline donors: effects of organic co-solvent and pH on reactions catalysed by Endo A and Endo M

The synthetic efficiency of endohexosaminidase-catalysed glycosylation reactions using N-glycan oxazolines as donors was investigated as two reaction parameters were varied. Both the addition of quantities of an organic co-solvent and modulation of reaction pH between 6.5 and 8.0 were found to have different effects on reactions catalysed by either Endo A (and two available mutants) or Endo M, indicating subtle differences between these two family GH85 enzymes. Fine tuning of reaction pH, or the addition of quantities of an organic co-solvent, resulted in beneficial increases in achievable synthetic efficiency by effecting a reduction in the rate of competitive hydrolytic processes.     

Three-dimensional structural resemblance between leucine aminopeptidase and carboxypeptidase A revealed by graph-theoretical techniques.

Using 3-D searching techniques based on algorithms derived from graph theory we have established a striking structural similarity between the structure of bovine carboxypeptidase A and that of the C-terminal domain of bovine leucine aminopeptidase. There is no significant sequence homology between the aminopeptidases and the carboxypeptidases but the strong structural relationship detected in this complex fold suggests that there may be a very remote divergent evolutionary relationship between these two enzyme classes.     

Nutritional Energy Stimulates NAD+ Production to Promote Tankyrase-Mediated PARsylation in Insulinoma Cells

The poly-ADP-ribosylation (PARsylation) activity of tankyrase (TNKS) regulates diverse physiological processes including energy metabolism and wnt/β-catenin signaling. This TNKS activity uses NAD⁺ as a co-substrate to post-translationally modify various acceptor proteins including TNKS itself. PARsylation by TNKS often tags the acceptors for ubiquitination and proteasomal degradation. Whether this TNKS activity is regulated by physiological changes in NAD⁺ levels or, more broadly, in cellular energy charge has not been investigated. Because the NAD⁺ biosynthetic enzyme nicotinamide phosphoribosyltransferase (NAMPT) in vitro is robustly potentiated by ATP, we hypothesized that nutritional energy might stimulate cellular NAMPT to produce NAD⁺ and thereby augment TNKS catalysis. Using insulin-secreting cells as a model, we showed that glucose indeed stimulates the autoPARsylation of TNKS and consequently its turnover by the ubiquitin-proteasomal system. This glucose effect on TNKS is mediated primarily by NAD⁺ since it is mirrored by the NAD⁺ precursor nicotinamide mononucleotide (NMN), and is blunted by the NAMPT inhibitor FK866. The TNKS-destabilizing effect of glucose is shared by other metabolic fuels including pyruvate and amino acids. NAD⁺ flux analysis showed that glucose and nutrients, by increasing ATP, stimulate NAMPT-mediated NAD⁺ production to expand NAD⁺ stores. Collectively our data uncover a metabolic pathway whereby nutritional energy augments NAD⁺ production to drive the PARsylating activity of TNKS, leading to autoPARsylation-dependent degradation of the TNKS protein. The modulation of TNKS catalytic activity and protein abundance by cellular energy charge could potentially impose a nutritional control on the many processes that TNKS regulates through PARsylation. More broadly, the stimulation of NAD⁺ production by ATP suggests that nutritional energy may enhance the functions of other NAD⁺-driven enzymes including sirtuins.